A Powerful Nonviral Vector for In Vivo Gene Transfer into the Adult Mammalian Brain: Polyethylenimine

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Abstract
Nonviral gene transfer into the central nervous system (CNS) offers the prospect of providing safe therapies for neurological disorders and manipulating gene expression for studying neuronal function. However, results reported so far have been disappointing. We show that the cationic polymer polyethylenimine (PEI) provides unprecedentedly high levels of transgene expression in the mature mouse brain. Three different preparations of PEI (25-, 50-, and 800-kD) were compared for their transfection efficiencies in the brains of adult mice. The highest levels of transfection were obtained with the 25-kD polymer. With this preparation, DNA/PEI complexes bearing mean ionic charge ratios closest to neutrality gave the best results. Under such conditions, and using a cytomegalovirus (CMV)–luciferase construction, we obtained up to 0.4 106 RLU/μg DNA (equivalent to 0.4 ng of luciferase), which is close to the values obtained using PEI to transfect neuronal cultures and the more easily transfected newborn mouse brain (106 RLU/μg DNA). Widespread expression (over 6 mm3) of marker (luciferase) or functional genes (bcl2) was obtained in neurons and glia after injection into the cerebral cortex, hippocampus, and hypothalamus. Transgene expression was found more than 3 months post-injection in cortical neurons. No morbidity was observed with any of the preparations used. Thus, PEI, a low-toxicity vector, appears to have potential for fundamental research and genetic therapy of the brain. Nonviral gene transfer into the adult mammalian brain offers the prospects of safe therapies and new experimental possibilities. However, results reported so far have been disappointing. We show that the cationic polymer polyethylenimine (PEI) provides high levels of transgene expression in the mature mouse brain. We used polymer preparations of low mean molecular weight and a polymer/DNA charge ratio near neutrality. Widespread expression (over 6 mm3) of marker (luciferase) or functional genes (bcl2) was obtained in neurons and glia after injection into the cerebral cortex, hippocampus, and hypothalamus. Transgene expression was found more than 3 months post-injection in cortical neurons. Thus, PEI, a low-toxicity vector, appears to have potential for fundamental research and genetic therapy of the brain.